THE CHEMISTRY OF THE PROTEINS 101 



esters pass over into the distillate, but they can be recovered by 

 acidifying with hydrochloric acid and evaporating, when they 

 are obtained as amino acid hydrochlorides. The dark brown 

 residue which remains in the distilling flask is fractionally 

 distilled in vacuo, leaving a mass consisting of tyrosine and 

 other amino acids, which in the first method are contained in 

 the mass of potassium carbonate. 



The distillation in vacuo is carried out first at the pressure 

 of 10-12 mm. produced by the water pump, and then at a 

 pressure of o - 5 mm. produced by a Geryck pump. At 10-12 mm., 

 three fractions (up to 6o°, 8o°, ioo°) are collected, the tempera- 

 ture taken being that of the water bath ; the distillation is 

 then continued at 0*5 mm., the first fraction being that at the 

 temperature of the boiling water bath ; several fractions are then 

 collected up to 160 , the temperature being procured by the use 

 of an oil bath. The residue, which does not distil at this 

 temperature, consists of a dark brown oil ; it sets to a hard 

 mass on cooling, and contains, amongst other substances, 

 leucinimide. 



The various fractions, so obtained, consist of a mixture of 

 the esters of two or more amino acids, and they are again 

 fractionated in vacuo ; this is not essential for the fractions dis- 

 tilling above ioo° C. at 05 mm. pressure, which contain the esters 

 of phenylalanine, aspartic acid, glutamic acid, and serine. These 

 esters are separated by taking advantage of their different 

 solubilities in water, ether, petroleum ether ; e.g. phenylalanine 

 ester is soluble with difficulty in water, and is extracted from 

 the aqueous solution of the esters with ether, the esters of 

 aspartic and glutamic acids remaining in the water. Serine 

 ester is insoluble in petroleum ether; on mixing the fraction 

 containing it with petroleum ether it separates as an oil, the 

 esters of the three other acids remaining in solution. 



Finally the esters must be converted into the amino acids ; 

 in the case of the lower-boiling fractions, they are hydrolysed 

 by boiling with water under a reflux condenser for five to six 

 hours until the alkaline reaction disappears ; leucine may 

 crystallise out during this process. The esters in the higher- 

 boiling fractions which contain glutamic and aspartic acids are 

 hydrolysed by boiling with baryta ; if hydrolysed by water, 

 they remain as acid esters. On cooling, aspartic acid may 

 separate as the barium salt of the racemised acid. The 



